Objective To establish interstitial cells of Cajal （ICC） loss models caused by incomplete small intestinal obstruction in rats with modified method and verify it. Methods Modified method was used to establish the models， making the ring around the small intestine but not through it. Morphological changes were observed by general signs， pathological changes were tested by HE staining and transmission electron microscope （TEM）， and changes of ICC number were tested by immunohistochemistry staining. Results Success rate of this method was 56% （28/50）， weight loss happened compared with before operation in ileus group （P＜0.01）. Hyperemia and swelling were observed in ileus group， and gastric retention was obvious. Results of HE staining and TEM showed that there was obvious inflammatory change， and ICC reduced was observed by immunohistochemistry. Conclusion ICC loss models caused by incomplete small intestinal obstruction meet the basic performance， and can be used for further research.
Abstract: Objective To induce calcification in aortic valvular interstitial cells (VICs) in vitro and observe the shift of cellular phenotype during the process. Methods Porcine aortic VICs were isolated and expanded by collagenase methods. Fluorescent staining was performed to identify the interstitial cells. VICs at 48 passages were used for experiments. The cells were divided into two groups: the experimental group in which cells were cultured in osteogenic media supplemented with βglycerophosphate, vitamin C and dexamethasone, and the control group in which cells were cultured in normal media. After 2 weeks, calcified nodules were quantified. Calcium deposit was stained and measured by Alizarin Red S staining and assay. Real time reverse transcription polymerase chain reaction (RTPCR) was performed to measure expression of alpha smooth muscle actin (α-SMA) and calcification related factors such as osteocalcin, osteopontin and Corebinding factor α1/Runx2 (Cbfα1/Runx2). Results VICs were successfully harvested from porcine aortic valves, identified by positive staining of α-SMA, vimentin and negative staining of Von Willebrand factor (vWF). VICs could calcify after 2 weeks of osteogenic induction with calcified nodules formed. Quantification of calcified nodules and calcification deposit were significantly higher (Plt;0.05) in the experimental group than those in the control group (156.25±17.38 vs. 2.50±1.29, 17.52±2.04 vs. 1.00±0.22). Real Time RT-PCR indicated that expression of α-SMA, as well as calcification related markers like osteocalcin, osteopontin and Cbfα1/Runx2 was much higher in the experimental group than those in the control group (Plt;0.05). Conclusion VICs are activated during the progress of calcification with phenotype shifting to contraction and ossification, which might be the pathological basis of valvular calcification.
目的 探讨新型玻璃化冷冻法对人卵巢组织超微结构的保存效果，尤其是对卵巢间质细胞的保存效果。 方法 收集2007年6月－2009年1月在我院行妇科手术的患者卵巢组织共8例，使用新型玻璃化冷冻法和慢速冷冻法保存，比较两种冷冻方法对于始基卵泡卵母细胞、颗粒细胞、卵泡周围间质细胞超微结构保存效果。 结果 对于始基卵泡卵母细胞和颗粒细胞，发现两种冷冻方法之间无显著差异（P＞0.05）；在间质细胞保存中，新型玻璃化冷冻法与慢速冷冻法相比较，正常间质细胞百分率增加（P＜0.05）。 结论 新型玻璃化冷冻法与慢速冷冻法相比能更好地保存卵泡周围间质细胞，在深低温保存人卵巢组织中具有较广阔的应用前景。
目的 初步观察电针足三里穴对不全肠梗阻大鼠小肠Cajal间质细胞（interstitial cells of Cajal，ICC）数量变化的影响，为进一步探讨电针足三里穴对ICC表型变化的影响奠定基础。方法 采用圈套法造成不全肠梗阻从而建立ICC数目减少的SD大鼠模型。取20只雌性大鼠采用简单随机法均分成正常对照组、不全肠梗阻30d未电针足三里穴组（梗阻组）、不全肠梗组30d电针足三里穴组（足三里组）和不全肠梗阻30d电针阴陵泉穴组（阴陵泉组） 4组。其中足三里组和阴陵泉组分别连续电针足三里穴或阴陵泉穴7d后，取小肠组织采用免疫组化方法以及免疫荧光观察ICC数量的变化。结果 正常对照组、足三里组、梗阻组及阴陵泉组ICC阳性面积分别为（102 051.00±16 969.38） μm2、（92 642.12±14 854.49） μm2、（45 221.33±6 230.20） μm2和（63 136.16±8 863.91） μm2，各组间差异有统计学意义（F=21.240，P＜0.001），其中足三里组的ICC阳性面积较梗阻组高（P＜0.05）。结论 电针足三里穴可使不全肠梗阻大鼠小肠受损的ICC数量得到部分恢复，但其具体机理有待进一步研究。
Objective To investigate the express of ERβ protein in female slow transit constipation (STC) patients. Methods Immunohistochemistry and Western blot technique were used to detect the distribution and expression of estrogen receptor β (ERβ) protein of 20 patients with STC and 20 aged-matched controls. Results ERβ expressions were detected in mucous layer, myenteric nerve plexus and submucous nerve plexus in two groups. In comparison with the control group, the expression of ERβ protein of STC group was much lower (Plt;0.01). The expression of ERβ protein of sigmoid colon in STC group was significantly lower than that in control group (Plt;0.05). Conclusion The expression of ERβ protein decreased in myenteric and submucous nerve plexus of sigmoid colon tissues may involve in the pathogenesis of STC.
Objective To study the effects of long term application of cathartics on electromyography of rat colon, and to explore the role of interstitial cells of Cajal (ICC) in it. Methods Colonic slow waves of the rat was examined after 3-month feeding of phenolphthalein, and ICC in myenteric plexus was observed by ZIO method, and ultrastructure changes of nerves and ICC was observed. Results The frequency of slow waves of cathartic colon was reduced significantly(Ｐ＜0.05). The distribution of ICC in myenteric plexus was uneven, and the processes were mussily connected each other. Vacuolar degeneration of axon and ICC-like cells was revealed by electron microscope in myenteric plexus of cathartic colon. Conclusion Long term application of phenolphthalein could reduce the frequency of colonic slow waves, and the possible mechanism was degeneration of ICC and myenteric plexus nerves.
ObjectiveTo investigate the changes of amount of interstitial cells of Cajal (ICC) and expression of stem cell factor (SCF)/c-Kit in the process of cathartic colon induced by emodin in mice. MethodsA modified cathartic colon mouse model was established. Seventy-two healthy male Kunming (KM) mice were randomly divided into the blank control group and sustained drug delivery group.Morphological changes of colon in mice were observed; frozen section immunofluorescence was used to observed changes of amount of ICC; serum concentrations of SCF were examined by ELISA; Western blot was employed for observation of expression of SCF/c-Kit in colon. ResultsAfter the mice model were completed, the weight of mouse, length and diameter of entire colon were all reduced compared with the blank control group. The amount of ICC appeared to decline in the beginning of the first 6 weeks with emodin used, and significant decreased in 10-12 weeks. The serum concentrations of SCF first began to decline in 4 weeks with emodin used, and significantly decreased in 6 weeks, and continued at a low level after 8 weeks. The expression of c-Kit in colon began to decline in 4 weeks with emodin used and significantly reduced after 8 weeks. Conciusions The amount of ICC appear to slowly decline in the beginning of the first 12 weeks with emodin used, and significant decrease after 12 weeks.The serum concentrations of SCF and expression of c-Kit in colon have the dynamic changes in the meanwhile, and the changes of SCF are earlier than that of c-Kit. The trend of amount ofICC may have a certain relationship with changes of SCF and c-Kit.
Objective To investigate the effect of different concentrations of raloxifene (RAL) on the proliferation and apoptosis of human aortic valve interstitial cells (AVICs) in vitro. Methods AVICs were isolated from human aortic valve by collagenase type Ⅱ, and cultured in different concentrations (0 nmol/L, 0.1 nmol/L, 1 nmol/L,10 nmol/L, 100 nmol/L and 1 000 nmol/L) of RAL. AVICs cultured in 0 nmol/L RAL were treated as the control group and those in other concentrations of RAL as the experiment groups. The proliferation and apoptosis of AVICs were evaluated by Cell Proliferation Assay (MTS assay) on day 0, 3, 5, 7 and 9. Flow cytometry was used to detect the cell cycle and apoptosis of AVICs on day 7. Results MTS results showed that the optical density value at 490 nm was much less in 10 nmol/L RAL and 100 nmol/L RAL groups (P＜0.05) on day 5, 7 and 9 than that in the control group. Flow cytometry results demonstrated that S-phase rate (P＜0.05) and cell apoptosis rate (P＜0.05) on day 7 were lower in the 10 nmol/L and 100 nmol/L RAL groups compared with the control group. Conclusion RAL with suitable concentration can inhibit proliferation and apoptosis of AVICs, which will lay an important foundation for further research of the role of RAL on heart valve diseases.
Objective To investigate the role and mechanism of peroxisome proliferator-activated receptor gamma coactivator 1α (PGC-1α) in the activation of aortic valve interstitial cells (AVICs) in aortic stenosis. Methods Isolating primary AVICs and stimulating their activation with transforming growth factor β1 (TGF-β1, 30 ng/mL), the expression of PGC-1α was detected. The activation of AVICs induced by TGF-β1 was observed after overexpression of PGC-1α by adenovirus or inhibition of PGC-1α function by GW9662. The possible downstream molecular mechanism of PGC-1α in AVICs activation was screened. Finally, the phenotype was further verified in primary human AVICs. Results The expression of PGC-1α decreased after the activation of AVICs induced by TGF-β1 (control group: 1.00±0.18; 24 h: 0.31±0.10; 48 h: 0.32±0.06; 72 h: 0.20±0.07; P<0.05). Specific overexpression of PGC-1α by adenovirus inhibited the activation of AVICs induced by TGF-β1 stimulation (periostin: 3.17±0.64 vs. 1.45±0.54, P<0.05; α-smooth muscle actin: 0.77±0.11 vs. 0.28±0.06, P<0.05). On the contrary, inhibition of PGC-1α function by GW9662 promoted the activation of AVICs (periostin: 2.20±0.68 vs. 7.99±2.50, P<0.05). Subsequently, it was found that PGC-1α might inhibit the activation of AVICs through downregulating the expression of calcium/calmodulin-dependent protein kinase (CAMK1δ) (0.97±0.04 vs. 0.74±0.11, P<0.05), and downregulating the expression of CAMK1δ alleviated the activation of AVICs (periostin: 1.76±0.11 vs. 0.99±0.20, P<0.05). The possible mechanism was that the activation of mammalian target of rapamycin (mTOR) signaling pathway was inhibited by reducing the accumulation of reactive oxygen species (ROS) (778.3±139.4 vs. 159.3±43.2, P<0.05). Finally, the protective effect of PGC-1α overexpression was verified in the activated phenotype of human AVICs (periostin: 2.73±0.53 vs. 1.63±0.14, P<0.05; connective tissue growth factor: 1.27±0.04 vs. 0.48±0.09, P<0.05). Conclusions The expression of PGC-1α significantly decreases during the activation of AVICs induced by TGF-β1. The overexpression of PGC-1α significantly inhibites the activation of AVICs, suggesting that PGC-1α plays a protective role in the activation of AVICs. The possible mechanism is that PGC-1α can inhibit the activation of CAMK1δ-ROS-mTOR pathway. In conclusion, interventions based on PGC-1α expression levels are new potential therapeutic targets for aortic stenosis.